It has previously been demonstrated that about 30% of healthy Caucasian subjects are "nonresponders" in assays of the mitogenic activity of monoclonal mouse IgG1 (mIgG1) anti-CD3 antibodies (e.g., anti-Leu 4 and UCHT-1), and that this unresponsiveness is due to lack of monocyte helper function. In an immunofluorescence assay with fluorescence-activated cell sorter analysis, we studied the binding of phycoerythrin-conjugated anti-Leu 4 to monocytes from responders and nonresponders. Interaction was observed with monocytes from responders only, and was blocked by a murine monoclonal antibody (IV.3) directed to an epitope on the 40-kDa low affinity Fc receptor (FcRII). This indicates that the interaction represents binding of the Fc part of phycoerythrin-conjugated anti-Leu 4 to FcRII on responder monocytes. Indirect immunofluorescence with antibody IV.3 demonstrated, however, that monocytes from both responders and nonresponders express similar levels of FcRII. Thus, nonresponder monocytes apparently express a variant FcRII which is unable to bind the Fc part of mIgG1 antibodies. The anti-FcRII antibody completely blocked anti-Leu 4-induced (but not OKT3 (mIgG2a)-induced) T cell proliferation in cultures of peripheral blood mononuclear cells from responders. The results provide direct evidence that monocytes from anti-Leu 4 responders, but not monocytes from anti-Leu 4 non-responders, are able to bind the Fc part of mIgG1 to FcRII, and that this interaction with FcRII is essential for the mitogenic activity of mIgG1 anti-CD3 antibodies.